tetano
Editor, Senior Moderator
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2014 Feb;30(2):117-20.
[Construction and identification of H5 subtype avian influenza vaccine candidate based on flagellin].
[Article in Chinese]
Wang J, Kang X, Jiao Y, Xiong D, Song L, Yang Y, Pan Z, Jiao X.
Author information
Abstract
Objective To develop a novel H5 subtype avian influenza vaccine candidate based on flagellin. Methods HA1-2-fljB was amplified through splicing hemagglutinin gene fragment HA1-2 of A/Goose/Jiangsu/1/2000 H5N1 and Salmonella typhimurium II phase flagellin fljB gene by overlap PCR, and then the gene was inserted to pET32a(+) to construct the recombinant plasmid pET32a-HA1-2-fljB. The recombinant plasmid was transformed into E.coli BL21 (DE3) and was induced with IPTG. The expression of the fusion protein was identified by SDS-PAGE and Western blotting. The biological activity of the fusion protein was evaluated by stimulating HEK293-TLR5 cells. Results The recombinant plasmid pET32a-HA1-2-fljB was constructed. The fusion protein, whose relative molecule weight was about 100 000, had a good immunoreactivity. TLR5 bioassay demonstrated the flagellin part of the fusion protein could be recognized by TLR5 of HEK293-TLR5 cell line to secrete a higher level of IL-8 when compared with HA1-2 protein (P<0.01). Conclusion The flagellin-based fusion protein HA1-2-fljB with TLR5 bioactivity was expressed correctly, and the results would establish a basis for the further research on avian influenza virus H5 vaccine.
PMID:
24491047
[PubMed - in process]
http://www.ncbi.nlm.nih.gov/pubmed/24491047
[Construction and identification of H5 subtype avian influenza vaccine candidate based on flagellin].
[Article in Chinese]
Wang J, Kang X, Jiao Y, Xiong D, Song L, Yang Y, Pan Z, Jiao X.
Author information
Abstract
Objective To develop a novel H5 subtype avian influenza vaccine candidate based on flagellin. Methods HA1-2-fljB was amplified through splicing hemagglutinin gene fragment HA1-2 of A/Goose/Jiangsu/1/2000 H5N1 and Salmonella typhimurium II phase flagellin fljB gene by overlap PCR, and then the gene was inserted to pET32a(+) to construct the recombinant plasmid pET32a-HA1-2-fljB. The recombinant plasmid was transformed into E.coli BL21 (DE3) and was induced with IPTG. The expression of the fusion protein was identified by SDS-PAGE and Western blotting. The biological activity of the fusion protein was evaluated by stimulating HEK293-TLR5 cells. Results The recombinant plasmid pET32a-HA1-2-fljB was constructed. The fusion protein, whose relative molecule weight was about 100 000, had a good immunoreactivity. TLR5 bioassay demonstrated the flagellin part of the fusion protein could be recognized by TLR5 of HEK293-TLR5 cell line to secrete a higher level of IL-8 when compared with HA1-2 protein (P<0.01). Conclusion The flagellin-based fusion protein HA1-2-fljB with TLR5 bioactivity was expressed correctly, and the results would establish a basis for the further research on avian influenza virus H5 vaccine.
PMID:
24491047
[PubMed - in process]
http://www.ncbi.nlm.nih.gov/pubmed/24491047